Project #2: Electrophysiologic Analysis of RIM Function in Presynaptic Plasticity Elucidating the molecular basis and physiological significance of synaptic plasticity will lead to a more sophisticated understanding of the neural circuit modifications which underlie experience-dependent olasticity in both health and disease. Much is known about the mechanisms of postsynaptic forms of long asting plasticity. By comparison, however, relatively little is known about the underlying mechanisms of long lasting forms of presynaptic plasticity. In this proposal we focus on understanding the synaptic functions of a class of presynaptic, active zone proteins, RIMs, because of their required involvement in a prominent form of presynaptic LTP and their additional roles in basal neuretransmitter release and short-term plasticity. RIMs have several protein binding domains that interact with key components of synaptic vesicles and active zones. In Aim 1, we will evaluate the physiologic significance of RIM's diverse protein interactions by attempting to rescue the synaptic abnormalities of autaptic cultured hippocampal neurons lacking RIMs with mutant RIMs that disrupt individual protein interactions. In Aim 2, we will evaluate the functional roles of several different RIM isoforms by examining synaptic function in autaptic cultured neurons in which these are absent or overexpressed. In Aim 3, using a knockin mouse model, we will test the functional significance of mutating a key serine residue (S413A) hypothesized to be required for presynaptic LTP by evaluating synaptic function in autaptic cultured neurons and acute hippocampal and cerebellar slices prepared from these mutant mice. In Aim 4, we will further characterize several features of presynaptic LTP in the cerebellum in the context of Project 4 which examines its postulated contribution to motor learning. Taken together, these studies will help elucidate the molecular basis of multiple forms of presynaptic plasticity and enable the generation of tools that will facilitate the examination of their functional significance at the behavioral level. By defining RIMs'molecular interactions that mediate synaptic plasticity and behavior, we will generate information that will be critical for targeting these proteins as needed for the treatment of a wide range of neuropsychiatric diseases.